Key Lab of Drug Metabolism and Pharmacokinetics, China Pharmaceutical University, Nanjing, Jiangsu, China.
Key Lab of Drug Metabolism and Pharmacokinetics, China Pharmaceutical University, Nanjing, Jiangsu, China.
J Pharm Biomed Anal. 2022 Jan 20;208:114437. doi: 10.1016/j.jpba.2021.114437. Epub 2021 Oct 22.
ZL-01 is a novel dual-prodrug which shows promise to be an antiviral candidate for hepatitis C virus. Here we have established a liquid chromatography tandem mass spectrometry (LC-MS/MS) method for simultaneous determination of ZL-01 and its four metabolites (M1, M7, M8, and M9) in rat plasma with special consideration of ex vivo ZL-01, M1, and M7 stability. Several factors affecting the stability were investigated. EDTA and citric acid solution (1 M) were added to plasma to maintain the stability of analytes. The protein-precipitation method was selected with acetonitrile containing sofosbuvir as internal standard (IS). Adequate separation of ZL-01 and its metabolites was achieved on XSelect HSS T3 (3.5 µm, 4.6 × 150 mm) column by a gradient-elution with a mobile phase consisting of 0.1% formic acid and acetonitrile at a flow rate of 0.5 mL/min. The detection was performed on a triple quadrupole tandem mass spectrometer by multiple reaction monitoring (MRM) mode to monitor the precursor-to-product ion transitions of m/z 599.2→418.5 for ZL-01, m/z 529.7→398.2 for M1, m/z 330.5→182.0 for M7, m/z 260.3→112.1 for M8, m/z 261.3→113.2 for M9 and m/z 530.4→243.4 for IS. The calibration curves exhibited good linearity (r＞0.997) for all components. The lower limit of quantitation (LLOQ) was in the range of 1-2 ng/mL. The intra-day and inter-day precisions (RSD) at three different levels were both less than 10.2% and the accuracies (RE) ranged from -3.7-7.6%. The matrix effect and extraction recovery of them ranged from 84% to 110.3% and 88.3-106.3%. This LC-MS/MS method for the simultaneous quantitation of ZL-01 and its metabolites was developed successfully and applied in the pharmacokinetic studies of these in rats. Pharmacokinetic results indicated ZL-01 would be metabolized rapidly and M8 might be the main metabolites after oral absorption.
ZL-01 是一种新型双重前药，有望成为丙型肝炎病毒的抗病毒候选药物。在这里，我们建立了一种液相色谱串联质谱（LC-MS/MS）方法，用于同时测定大鼠血浆中的 ZL-01 及其四种代谢物（M1、M7、M8 和 M9），特别考虑了离体 ZL-01、M1 和 M7 的稳定性。研究了影响稳定性的几个因素。向血浆中加入乙二胺四乙酸和柠檬酸溶液（1 M）以维持分析物的稳定性。选择乙腈沉淀法，其中含有索非布韦作为内标（IS）。在 XSelect HSS T3（3.5 µm，4.6 × 150 mm）柱上，通过梯度洗脱，以含有 0.1%甲酸和乙腈的流动相在 0.5 mL/min 的流速下实现 ZL-01 和其代谢物的充分分离。通过多反应监测（MRM）模式在三重四极杆串联质谱仪上进行检测，以监测 m/z 599.2→418.5 的前体到产物离子跃迁，用于 ZL-01，m/z 529.7→398.2 用于 M1，m/z 330.5→182.0 用于 M7，m/z 260.3→112.1 用于 M8，m/z 261.3→113.2 用于 M9 和 m/z 530.4→243.4 用于 IS。所有成分的校准曲线均表现出良好的线性（r＞0.997）。定量下限（LLOQ）范围为 1-2 ng/mL。三个不同水平的日内和日间精密度（RSD）均小于 10.2%，准确度（RE）范围为-3.7-7.6%。它们的基质效应和提取回收率范围为 84%-110.3%和 88.3-106.3%。成功建立了同时定量测定 ZL-01 及其代谢物的 LC-MS/MS 方法，并应用于大鼠体内的药代动力学研究。药代动力学结果表明，ZL-01 会迅速代谢，M8 可能是口服吸收后的主要代谢物。
